The proposed study is intended to elucidate some of the synthetic mechanisms involved in E. coli DNA replication by examining conditional mutants which are deficient in specific components of the replication machinery. Three investigatory approaches are planned: 1) Mutant extracts, from dnaB strains, immobilized on cellophane membranes will be allowed to synthesize DNA in vitro, both alone and as complemented by other extracts. The size of the DNA produced will be analyzed by alkaline sucrose gradient centrifugation. Parallel in vivo analysis will also be done. Intragenic complementation of dnaB mutants will also be analyzed in heteroallelic diploids by measuring the alteration of the GroP minus phenotype which affects lambda bacteriophage growth. 2) By using double dna mutants, the construction of a DNA synthesis protein interaction map will be attempted, again using the alteration of the GroP minus phenotype as a measurement. 3) Finally, a search for new dna genes will be attempted by the isolation of more mutants using a new technique. It is hoped and expected that this work will eventually make it possible to conduct similar studies in eukaryotes where it should be of manifold importance in the study of differentiation and tumor cell growth.